A shared mucosal gut microbiota signature in primary sclerosing cholangitis before and after liver transplantation

Abstract

Background and aims: Several characteristic features of the fecal microbiota have been described in primary sclerosing cholangitis (PSC), whereas data on mucosal microbiota are less consistent. We aimed to use a large colonoscopy cohort to investigate key knowledge gaps, including the role of gut microbiota in PSC with inflammatory bowel disease (IBD), the effect of liver transplantation (LT), and whether recurrent PSC (rPSC) may be used to define consistent microbiota features in PSC irrespective of LT.

Approach and results: We included 84 PSC and 51 liver transplanted PSC patients (PSC-LT) and 40 healthy controls (HCs) and performed sequencing of the 16S ribosomal RNA gene (V3-V4) from ileocolonic biopsies. Intraindividual microbial diversity was reduced in both PSC and PSC-LT versus HCs. An expansion of Proteobacteria was more pronounced in PSC-LT (up to 19% relative abundance) than in PSC (up to 11%) and HCs (up to 8%; Q FDR < 0.05). When investigating PSC before (PSC vs. HC) and after LT (rPSC vs. no-rPSC), increased variability (dispersion) in the PSC group was found. Five genera were associated with PSC before and after LT. A dysbiosis index calculated from the five genera, and the presence of the potential pathobiont, Klebsiella , were associated with reduced LT-free survival. Concomitant IBD was associated with reduced Akkermansia .

Conclusions: Consistent mucosal microbiota features associated with PSC, PSC-IBD, and disease severity, irrespective of LT status, highlight the usefulness of investigating PSC and rPSC in parallel, and suggest that the impact of gut microbiota on posttransplant liver health should be investigated further.

Graphical abstract

FIGURE 1

Study flowchart showing main groups and analyses performed. In a first step, microbiota analysis of the main study cohorts was performed. Next, identification of overlapping microbiota features between PSC and rPSC was performed. Last, identified microbiota features were used to investigate clinical correlates, among them LT‐free survival and time to diagnosis of recurrent PSC (recurrence‐free survival).

FIGURE 2

Diversity metrics for PSC, PSC‐LT, and HCs. (A) Principal coordinate analysis plot (Bray–Curtis dissimilarity) showing that PSC and PSC‐LT have unequal microbiota composition (PERMANOVA: R ²: 0.017, pseudo‐F: 7.38, p = 0.001; PERMDISP: p = 0.74), and that both are unequal compared with HCs, which, in part, is explained by higher dispersion in PSC and PSC‐LT (PERMANOVA: R ²: 0.041, pseudo‐F: 15.60, p = 0.001 and R ²: 0.064, pseudo‐F: 19.72, p = 0.001, respectively, and PERMDISP: p = 0.001 and p = 0.001, respectively). (B) Principal coordinate analysis in panel A colored according to alpha diversity (observed ASVs), showing that the pattern along the PCA1‐axis is driven by alpha diversity, Spearman correlation coefficient: −0.65, p < 2.2 × 10⁻¹⁶. (C) Reduced alpha diversity (Shannon diversity index) in PSC and PSC‐LT compared with HC in all segments and similar alpha diversity between segments in all groups. (D) Reduced alpha diversity (observed ASVs) in all available segments in PSC and PSC‐LT compared with HCs. PC, principal coordinate; PSC‐LT, transplanted PSC.

FIGURE 3

Phylum distribution in study cohorts and taxonomic heatmaps in PSC versus PSC‐LT. (A) Mean relative abundance of major phyla in HC, PSC, and PSC‐LT. ✧ Increased in PSC‐LT versus PSC. ✢ Reduced in PSC versus HC. ▲ Increased in PSC‐LT versus HC. ✖ Reduced in PSC‐LT versus HC. Significance‐level Q_FDR < 0.05. (B) Log₂‐fold change of significantly increased or decreased genera (Mann–Whitney U test) in PSC compared with PSC‐LT. Log₂‐fold change >0 indicates increased relative abundance in PSC. *p < 0.05; ****Q_FDR < 0.05. PSC‐LT, transplanted PSC.

FIGURE 4

Diversity metrics and taxonomic heatmap in PSC with concomitant IBD. (A) Principal coordinate analysis plot (Bray–Curtis dissimilarity) showing that PSC patients (irrespective of LT status) with IBD have significantly different microbiota composition compared with PSC without IBD (PERMANOVA: R ²: 0.0082, pseudo‐F: 3.61, p = 0.001; PERMDISP: p = 0.092). (B) No significant differences in alpha diversity (Shannon diversity index) between PSC with IBD and PSC without IBD (irrespective of LT status). (C) Alpha diversity (Shannon diversity index) in PSC‐LT with IBD compared to PSC‐LT without IBD. (D) Taxonomic heatmap of significantly increased or decreased genera (Mann–Whitney U test) in PSC‐LT and PSC with IBD compared to those without. Log₂‐fold change >0 indicates increased genus in PSC with concomitant IBD. *p < 0.05; ****Q_FDR < 0.05. PC, principal coordinate; PSC‐LT, transplanted PSC.

FIGURE 5

Diversity and overlapping genera in PSC and rPSC. (A) Reduced alpha diversity (Shannon diversity index) in rPSC (at inclusion) compared with HC. (B) Principal coordinate analysis plot (Bray–Curtis dissimilarity) showing that rPSC have significantly different dispersion compared to no‐rPSC (PERMANOVA: R ²: 0.02 pseudo‐F: 3.4, p = 0.001; PERMDISP: p = 0.006). (C) Five significantly increased or decreased genera associated with PSC or rPSC compared with HCs and no‐rPSC, respectively (Mann–Whitney U test). Log₂‐fold change >0 indicates increased genus in PSC/rPSC. Lachnospiraceae CAG‐56 (underscored) was, on average, increased in the ascending colon in PSC relative to HCs, although the median in PSC was lower than that of HCs. *p < 0.05; ****Q_FDR < 0.05. no‐rPSC, transplanted PSC without rPSC; PC, principal coordinate.

FIGURE 6

Microbiota associations with disease severity and survival. (A) LT‐free survival was reduced in those with a dysbiosis index (sigmoid colon) above median. RMST 5.52 versus 8.99 years, log‐rank: p = 0.05, Kaplan–Meier plot. (B) Recurrence‐free survival from date of inclusion was reduced in those with dysbiosis index (sigmoid colon) above median. RMST 6.56 vs. 8.53 years, p = 0.02, Kaplan–Meier plot. (C) Mayo PSC risk score and FIB‐4 score were increased in PSC patients (regardless of transplantation status) positive for Klebsiella in gut mucosal samples. (D) LT‐free survival was reduced in patients positive for mucosal Klebsiella, in any segment, RMST 4.28 versus 7.87 years, p = 0.01, Kaplan–Meier plot. no‐rPSC, transplanted PSC without rPSC.